The present invention relates to an optically accessible microfluidic device, to a method for manufacture thereof, and to a system that uses the microfluidic diagnostic device.
As is known, devices of a micromechanical or microelectromechanical (MEMS) type are widely used in industry, medicine and research. MEMS devices offer the advantages of improving the reliability of the assay, reducing sample volumes, as well as reducing the time required for said activities, thus reducing the corresponding costs and improving efficiencies.
Diagnostic devices of a known type basically comprise a solid substrate, generally of a flat type, on which particular receptors, such as, for example, biomolecules (DNA, RNA, proteins, antigens, antibodies, haptens, sugars etc.) or chemicals, or microorganisms or parts thereof (bacteria, viruses, spores, cells, organelles, etc.) are grafted.
By “receptor” what is meant herein is any member of a binding pair or multiple, such that the receptor will bind to or react with, and thus detect, its binding partner(s). Thus, receptor includes traditional receptors such as protein receptor and ligand, but also any member of a binding or interacting multiple such as lectin, carbohydrates, streptavidin, biotin, protein, substrate, oligonucleotides, nucleic acid, porphyrins, metal ions, antibodies, antigens, and the like.
When these receptors come in direct contact with a sample to be analysed, the presence in said sample of molecules capable of binding or interacting with the receptor is detected in some way. For example, binding can be detected with fluorescent or phosphorescent labels, which, when excited with light radiation at a certain wavelength λe, emit a light radiation having a wavelength λf different from the wavelength λe.
Known fluorescence diagnostic devices comprise a compatible layer, the surface of which is functionalized in such a way as to present sensing areas comprising receptors provided with specific labels. The labels are activated (i.e., they emit a light at a wavelength λf) only when sample of molecules bind or interact with the receptors.
There are many different ways to set up tests involving optical signals. For example, a common three component binding assay uses a first antibody bound to a solid substrate which can bind to an antigen present in a sample solution. Antigen binding is then detected with a second antibody that binds to a different epitope of the same antigen and which has a fluorescent label attached thereto. Thus, the amount of fluorescence detected correlates with the amount of antigen in the sample.
Another example involves binding an oligonucleotide probe to the substrate (or free in solution), which then hybridizes to complementary DNA or cDNA or mRNA in the sample, and the double stranded nucleic acid can be detected with an intercalating dye, such as ethidium bromide, which fluoresces on exposure to UV light.
In yet another example, two fluorescent markers are brought in close proximity in the assay, and quenching of one marker is measured in fluorescence resonance energy transfer (FRET) based assays. The reverse is also possible, that is where the assay separates two labels allowing an increase in signal.
As yet another example, heavy metal binding to fluorophores can also be detected with fluorescent dyes. Regardless of the assay particulars, similar devices can generally be employed with optical assays.
The light radiation in assays such as these can then be collected by an appropriate detector, such as, for example, a photo-detector of a charge-coupled device (CCD) type or of a CMOS type, compatible with the wavelength λf of the emitted light radiation. The variation of light intensity is a function of the amount of specific labels activated or detected in the assay, and hence of the amount of target in the sample.
The aim of the present invention is to provide an optically accessible microfluidic diagnostic device, a method for manufacture thereof, and a diagnostic system that uses the microfluidic diagnostic device.
By “diagnostic” device herein, we do not imply that the device is used only for medical purposes, but rather a general purpose device that can diagnose the presence (or absence) or concentration of a particular analyte is intended.